A simple procedure is described for the isolation of protoplasts from plant cells using a high concentration of magnesium sulfate as an osmotic stabilizer. This new procedure was developed with callus cells of Symphytum officinale as a starting material, but was also successfully applied to other tissues and other plants. The floating protoplast fraction obtained had a purity of more than 90% and was highly enriched in nucleated protoplasts (about 80%). Microdensitometry of Feulgenstained fixed protoplasts from callus cells of Symphytum officinale revealed two ploidy levels (diploid and tetraploid), in contrast to protoplasts from leaf material that appeared to be diploid only.